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KMID : 0604220060130010005
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Korean Journal Investigative Dermatology
2006 Volume.13 No. 1 p.5 ~ p.12
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Expression of Human β-Defensin 2 in Keratinocytes and Inflammatory Skin Disease
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Ahn Ji-Young
Seo Seong-Jun
Hong Chang-Kwun
Jeong Mi-Sook
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Abstract
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Background: Defensins are major antimicrobial peptides(AMPs), small cationic, cystein rich peptides with a wide range of antimicrobial activity against gram positive and gram negative bacteria, yeasts, fungi, and virus. They may immediately respond to intruded microbes for prevention of further invasion. Although they are negligible in normal non-inflamed skin, their expression can be markedly enhanced with some conditioned stimulations. Atopic dermatitis and psoriasis are common inflammatory skin disease but frequent skin infections are observed in atopic dermatitis patients, but not in psoriasis patients.
Objective: Current research was done to identify the expressions of human β-defensins 2(HBD-2) in normal human keratinocyte(NHK) and HaCaT cells, and also to understand of the differences in susceptibility to infection between atopic dermatitis and psoriasis.
Methods: NHK and HaCaT cells were processed with UVB irradiation and lipopolysaccharide(LPS). Expressions of HBD-2 in NHKs and HaCaT cells were evaluated by using reverse transcription polymerase chain reaction(RT-PCR), Western blotting, and immunohistochemical staining(IHC) at 6, 12, and 24 hours post stimulation. Also expressions of HBD-2 in skin biopsy specimens from four atopic dermatitis patients and four psoriasis patients were checked by RT-PCR, Western blotting, and IHC.
Results: With time-sequential analyses of AMP mRNA expression by RT-PCR, the expression level of HBD-2 at 12 hours was the peak point of maximum expression. In both NHKs and HaCaT cells, LPS supplementation of 2.5μg/ml concentration was the most powerful stimulus compared with other stimuli such as 5.0μg/ml LPS treatment and UVB irradiation(20mJ/cm2). Western blotting of HBD-2 protein in NHKs and HaCaT cells, the peak expression of HBD-2 protein was also obtained with 2.5μg LPS stimulation at 12 hours post stimulation. Through IHC analyses, we were able to confirm that HBD-2 protein syntheses were markedly increased by stimuli such as UVB irradiation and LPS supplemenation in both NHKs and HaCaT cells, and in status of cell differentiation. RT-PCR did not reveal HBD-2 expression in the epidermis of skin from atopic dermatitis, but profound HBD-2 expression was observed in the upper portion of psoriatic epidermis. Western analyses and IHC confirmed the levels of protein expression in psoriatic epidermis.
Conclusion: Our results demonstrate that the presence of HBD-2 in NHKs and HaCaT cells, which were upregulated by UVB irradiation and LPS treatment. Therefore these HBD-2 may contribute to the host defense mechanism against various environmental insults including Gram negative microbes and UVB radiation. And HBD-2 may, in part, be responsible for the frequent skin infections observed in atopic dermatitis patients, but not in psoriasis patients.
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KEYWORD
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Human β defensin-2, Keratinocyte, HaCaT cell, Atopic dermatitis, Psoriasis
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